Our long-term goal is to develop treatments that may improve resilience to pathogen damage in patients with serious infections, including chronic and relapsing bacterial infections, for which effective treatments are lacking. Bacterial quorum sensing (QS) signals are important mediators of immunomodulation. However, it remains unclear how microbes utilize immunomodulatory signals to maintain a long-term presence and thwart clearance and how hosts avoid harmful inflammatory response to a pathogen?s presence. In our previous work, we identified a small volatile QS molecule excreted by Pseudomonas aeruginosa (PA), 2-aminoacetophenone (2-AA), that acts as an inter-kingdom ?infochemical? to alter immune responses and metabolism in a manner that trains the host to increase host tolerance/resilience (HT/R), which is defined as the host's ability to cope with bacterial encounter without a consequent reduction in fitness; these alterations allow the pathogen to avoid elimination and persist in mammalian tissues. The specific goal of this application is to decipher paradigmatically the mode of action of a bacterial product on host metabolome and epigenome in order to gain insights into the first mechanistic example of a QS molecule that reprograms the host metabolome and epigenome to promote HT/R. Our hypothesis is that the long-lasting immunomodulatory changes exerted by 2-AA through the sustained activity of histone deacetylase 1 (HDAC1) derive from 2-AA?promoted metabolic alterations; and that the interplay between host metabolome and epigenome contribute to the mutual bacterial-host fitness that defines HT/R. We propose to achieve our goal through experiments employing PA, a recalcitrant Gram-negative ESKAPE bacterium that defies eradication by antibiotics, forms biofilms, and exemplifies current clinically problematic pathogens. In Aim 1, we will examine how 2-AA maintains epigenetic reprogramming by assessing the molecular mechanisms of long-term immunomodulation in immune memory in vivo promoted by 2-AA tolerization, and determine the occurring metabolic changes by performing functional studies. In Aim 2, we will interogate the capacity of 2-AA to regulate macrophage autophagy in tissues and conduct a meticulous analysis of the key host players and cellular mechanisms that permit bacterial persistence. In Aim 3, using murine in vitro and in vivo systems in addition to human primary macrophages, we will evaluate the interplay between 2-AA promoted epigenetic control and metabolic alterations by undertaking molecular and functional approaches. The impact of these alterations in bacterial persitence, histone acetylation, energy metabolism and the efficacy of our novel anti-2AA compound will be asssessed using clinically relevant animal models of infection. In Aim 4, we will seek to identify 2-AA?s direct target using click chemistry. The proposed elucidation of ?host tolerance training? mechanisms is intended to improve our understanding of HT/R that remains extremely limited. In this context, the findings may offer opportunities for being applied to the development of therapeutic treatments that can train the host to augment resilience to infection. In principle, the knowledge to be obtained could be applicable to persistent infections triggered also by other bacteria because QS is broadly conserved among Gram-negative and Gram-positive human bacterial pathogens and, as such, 2-AA-like molecules are likely to exist in other bacteria.